1. Field of the Invention
The present invention relates to stable immunogenic products comprising immunogenic protein heterocomplexes for obtaining a humoral immune response with production of specific antibodies raised against one ore more antigens, in particular against a “self” antigen, as well as their use in the field of vaccines.
2. Related Art
Obtaining a high level antibody response from a given antibody, in an individual, is an object commonly sought, whether the antigen is a “foreign” antigen or a “self” antigen.
However, the problem of a good recognition of the antigen against which an antibody response is being sought, in an individual, should be solved in a number of cases, more particularly (a) when the antigen of interest behaves like a “hapten”, i.e. a low molecular mass chemical structure which is little or not immunogenic under a free form, but that, once fixed on a high molecular mass molecule, is able to induce the production of specific antibodies of such a hapten, and (b) when the antigen of interest is a self protein, i.e. a protein being naturally produced in the individual, for which there exists an immune tolerance due to the deletion of corresponding lymphocyte T clones, during the development of the immune system.
In order to cause, or increase, the recognition of an antigen of interest by B cells, various immunogenic constructions were developed in the state of the art.
A first immunogenic construction form comprises a covalent coupling of the antigen of interest on a carrier molecule, the carrier molecule bringing structures recognized by the auxiliary T lymphocytes (“T helper” cells), in association with class II molecules of the Histocompatibility Major Complex (HMC), and activating the auxiliary T lymphocytes then producing various cytokins, amongst which IL-2, said cytokins activating in turn the specific B cell clones of the antigen of interest. The specific B cells of the antigen of interest, once activated, multiply and produce antibodies specific to the antigen of interest, which is the objective being sought. Generally, such a type of immunogenic constructions comprises products of the covalent chemical coupling between the antigen of interest and the carrier molecule, which, after purification and removal steps of the non coupled products, are final products with a well defined chemical structure.
The first form of an immunogenic construction is for example illustrated by the article by Richard and al. describing the preparation of products of the covalent coupling between IL-9 and ovalbumin (Proc. Natl. Acad. Sci. USA, Vol. 97(2): 767-772). It is also illustrated in such U.S. Pat. No. 6,340,461 (Terman) which discloses coupling products between one or more copies of an antigen of interest, against which a specific antibody response is being sought in an individual, and a carrier molecule consisting in a “Superantigen”. The antigen of interest is coupled exclusively covalently to the carrier molecule, for example, by means of glutaraldehyde (also called “pentanedial”), the non-covalently coupled products being removed in order to obtain a chemically well-defined final product.
Optionally, the product from the covalent coupling between the antigen of interest and the superantigen could be prepared in the form of a polymer of said coupling product, for example, through a non covalent bond of the monomeric coupling products between one another, through ionic interactions, adsorption interactions as well as biospecific interactions. For example, the monomeric coupling products could form complexes with highly positively or negatively charged molecules, through salt bridges produced in low ionic strength conditions. Larges complexes of monomeric coupling products are prepared using charged polymers such as poly(L-glutamic acid) or poly(L-lysine) polymers. According to another embodiment of a monomeric coupling product polymer, the exclusively covalent coupling products between the antigen of interest and the superantigen could be adsorbed or coupled non covalently at the surface of microparticles, such as latex beads or other hydrophobic polymers.
A second embodiment of such immunogenic constructions commonly called “MAP” structure (for “Multi-Antigenic Protein”) generally have the form of a protein backbone comprised of a linear or branched, poly(lysine) polymer, onto which one or more antigens of interest are covalently bound.
A third embodiment of such immunogenic constructions consists in microparticles onto which fixed the antigen(s) of interest is/are bound. Various forms of antigen carrier microparticles are known.
For example, iscomes (for “immunostimulating complexes”) are known comprised of an antigenic complex and an adjuvant, the QuilA compound.
Liposomes are also known having the same drawbacks as the iscomes, i.e. more particularly some toxicity and immunological side effects, due to their lack of purity.
Biodegradable microparticles are also known such as lactic acid and glutamic acid polymers (Aguado and Lambert, 1992, Immuno. Biol., Vol. 184: 113-125) as well as starch particles (U.S. Patent Application 2002/0098203—Gutavsson et al.), in the polymeric matrix of which antigens of interest are trapped. Such particles release the antigen under their soluble form during the degradation of the polymeric matrix.
Particles have also been disclosed exclusively comprised of hybrid recombinant proteins, as disclosed in French Patent Application FR 2,635,532 (Thiollais et al.).
Porous microspheres are also known wherein the antigens are immobilized within micropores through captation or physical coupling, as disclosed in the U.S. Pat. No. 5,008,116 (Cahn).
However, the various solutions suggested in the state of the art share in common at least one technical inconvenient related to their preparation method, i.e. the loss of a high proportion of the antigenic material of interest, due to a necessary step for removing the non coupled or non adsorbed antigens.
Moreover, while the prior art techniques allow to provide an association between the low molecular mass antigen of interest with a carrier molecule, they are generally not adapted to coupling a high molecular mass antigen of interest, for example, of more than 10 kDa, with the carrier molecule, because, in particular, of steric hindrances preventing coupling a high number of molecules of antigens of interest having a high molecular mass with an identical carrier molecule.
Finally, most if not all the known peptide antigenic constructions encompass in their structure a single carrier molecule, which is a technical inconvenient when the objective is to induce a preventive or therapeutic immune response both against the antigen of interest and the carrier molecule itself.
There is therefore a need in the state of the art for improved immunogenic constructions allowing for the production of a high level of antibodies specific to an antigen of interest in an individual where such a humoral immune response is sought, being less expensive, simple to prepare and able to be synthesized reproducibly.